1 x G-VDTP 60,000ng in 30ml

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SKU: 1xGVDTP_60,000 Kategorija:

Opis

G-VDTP for research purposes only.

Product name: G-VDTP

Identified uses: Research purposes only. Not for human consumption.

Format: 30ml amber vials

Sterility test: Sterility/Endotoxin tests according to Pharmacopeia 8.0 (Eur) using “Steritest” and “LAL”

Activity tests: Cancer cell viability reduced by >40% in 72hrs & Reactive oxygen species generation (macrophage activation)

Contents: 60,000ng Vitamin D Transport Protein (VDTP), 330ng Oleic Acid suspended in 30ml sterile natrium chloratum (saline solution)

Classification: Classified in the trade tariff under commodity codes  21 06 10 80 70 &  29 42 00 00 90

VDTP weight: 51,423 Da

PH: pH7

Colour: Colourless

Clarity: Clear

Preparation: Prepared in an ISO & GMP registered laboratory in the EU..

Format: Amber glass vials with crimp cap.

Presentation: General appearance of bottle and label should be satisfactory. Label data should be correct.

Storage: Can be frozen for long term storage – use within 18 months. Otherwise keep refrigerated and use within 3 months. Avoid light direct sunlight. Shake well before use. Keep away from children.

Registration: UK & EU Registered food supplement research products.

Download full specification sheet here.

CYTONIC DODATAK PREHRANI – ANALITIČKI PARAMETR

NAZIV PROIZVODA:

Version: final (audited)

NAZIV PROIZVODA: Cytonic dodatak prehrani

1. Formulacija i karakteristike 

Formulacija i karakteristični organoleptički parametri su potvrđeni.

2. Koncentracija proteina

Koncentracija proteina u neprerađenom pripravku VDTP određena je Bradfordovim testom (IB-16.05.00).

Nakon obrade pripremljene su konačne otopine VDTP u koncentracijama:

167 ng / ml; 1.000 ng / mI i 3.334 ng / mI (± 10%) u odnosu na početnu koncentraciju.

3. Identitet proteina potvrđen je masenom spektrometrijom

Točnost mjerenja je ± 1%, stoga je rezultat od 50.969Da prihvatljiv za VDTP (51.243Da).

4. Čistoća

Čistoća proteina procijenjena je denzitometrijom.

Količine 0,5, 1 i 2 ktg VDTP proteina napunjene su na SDS-PAGE gel. Nakon elektroforetskog odvajanja, gel je obojen Coomassie briljantnim plavim. Analiza intenziteta apsorpcije provedena je pomoću softvera Image Lab 5.2.1.

5. Razina endotoksina

Razina endotoksina u VDTP procijenjena je primjenom testa Limulus Amebocyte Lysate (LAL). 

6. Sterilnost

Ukupni aerobni broj mikroba procijenjen je inokulacijom Trypone sojinog agara sa 100 [..t1 VDTP i inkubacijom na 37 ° C tijekom 5 dana. Nisu primijećene kolonije.

Ukupni kombinirani broj kvasca i plijesni procijenjen je inokulacijom Sabourauda (+ dekstroza + kloramfenikol) sa 100 µl VDTP i inkubacijom na 37 ° C tijekom 5 dana. Nisu primijećene kolonije.

Oba su ispitivanja rađena u tri primjerka.

Odsutnost E.coli u 10 g potvrđena je cijepljenjem McConkey juhe s 10 g VDTP i inkubacijom na 37 ° C tijekom 5 dana. Nije primijećena zamućenost.

Referenca: USP29-NF24 stranica 3087 http://www.pharmacopeia.cn/v29240/usp29nf24s0_c2023.html#usp29n124s0_c2023thNovember 2016]

7. Mikoplazma

Prisutnost mikoplazme u VDTP procijenio je NAT (Tehnike pojačavanja nukleinskih kiselina). Rezultat je bio negativan. 

8. Retrovirus

Prisutnost retrovirusa u VDTP procijenjena je pomoću PERT-a (Test poboljšane reverzne transkriptaze proizvoda). Rezultat je bio negativan. 

9. pH

pH otopine VDTP određen je pomoću pH-metra Hanna Instruments HI2212. Mjerenje je ponovljeno tri (3) puta i izračunata je prosječna vrijednost:

Anticancer efficacy of Vitamin D Transport Protein (VDTP) can be ascribed at least to three biological properties of the molecule:

1) activation of tumoricidal macrophages;

2) inhibition of tumor-induced angiogenesis and

3) direct inhibition of cancer cell proliferation, migration and metastatic potential.

Reference: Pacini S, Punzi T, Morucci G, Gulisano M and Ruggiero M: Effects of vitamin D-binding protein-derived macrophage activating factor on human breast cancer cells. Anticancer Res 32: 45-52, 2012

In order to test whether Cytonic proteins (CP) inhibits cancer cells proliferation, we treated MCF-7 cells (cell line derived from human breast cancer) with various concentrations of proteins. 

It has been showed that Cytonic proteins directly inhibits breast cancer cells proliferation in dose- dependent manner.

Results

Fig. 1: Breast cancer cells viability after treatment with various concentration of Cytonic proteins (CP). After 72h incubation with Cytonic proteins viability reduction in dose-dependent manner was observed. The effect was the most significant for 400 ng/ml, where cell viability is reduced to ≈ 59.6% with respect to control cells.

Inactive and Active Immune Cells

Immature immune cells are created in our bone marrow, lymph glands and spleen. Then they are released into our blood plasma where they circulate with the blood looking for a specific protein to become fully active and functional immune cells. The protein required to activate all of our immune cells is Vitamin D Transport Protein (VDTP)

In the first 10 seconds of the above video you will see an ‘inactive’ immune cell, it is consuming just enough energy to stay alive but cannot function as a mature immune cells until it is activated.

From 10 seconds onwards you will see different types of active immune cells in action, despite their differences they carry out similar functions and that is to ‘seek and destroy’. We can see the active immune cells using their pseudopods to check red blood cells for any proteins which may indicate an infection. If a ‘suspect’ cell is discovered the the immune cell will consume it then use oxidise bursts to breakdown the infected cell into amino acids and proteins which are recycled into new and healthy red cells.

At 36 seconds we observe an immune cell homing in an a ‘Helmut Cell’ half a red blood cell that served no purpose in the body and therefore must be eliminated and recycled.

ACTIVITY REPORT OF OMNIA & CYTONIC FOOD SUPPLEMENTS.

Macrophages are key immune effector cells. They display remarkable plasticity and can change their physiology in response to environmental stimuli, giving rise to different populations of cells with distinct functions. Depending on circumstances, activation of macrophages can result in enhanced microbicidal or tumoricidal capacity, wound-healing activity or differentiation into regulatory macrophages’.

Depending on circumstances, activation of macrophages can result in enhanced microbicidal or tumoricidal capacity, wound-healing activity or differentiation into regulatory macrophages’. 

Vitamin D Transport Protein has been shown to play a role in inflammation by activating macrophages and inducing oxidative burst. Oxygen-dependent mechanisms are known to mediate the anti-microbial and anti-tumor activities of macrophages2.

OBJECTIVE

The aim of this experiment was to evaluate whether Cytonic and Omnia compounds displayed immune activity measured upon oxidative stress caused by ROS (reactive oxygen species) generation.

SAMPLES

  • Omnia Drops (10, 100 and 1000-fold dilutions)
  • Chondroitin sulfate (2.5, 0.25 and 0.025 mg/ml) corresponding to concentrations present in respective dilutions of Omnia Drops
  • Oleic acid (1.25, 0.125, 0.0125 mg/ml) corresponding to concentrations present in respective dilutions of Omnia Drops
  • Oleic acid (1.25, 0.125, 0.0125 mg/ml), dissolved in ethanol (final ethanol concentration in the medium C=0.05%).
  • Cytonic protein (3.3, 0.33, 0.033 p.g/m1)

*All dilutions were prepared in PBS buffer, unless indicated otherwise. 

METHOD

Immortalized bone marrow-derived macrophages (BMDM) and murine macrophage-like cell line RAW 246.7 were used as test cells. ROS production was measured with CeIIROX® Green (CRG) probe. CRG exhibits bright green fluorescence upon oxidation by ROS and subsequent binding to DNA, therefore enables oxidative burst measurement.

Cells were stained with CRG and incubated with analyzed compounds, PMA (positive control; ROS inducer) or buffer only (negative control).

After incubation cells were fixed in formaldehyde and stained with DAPI to visualize nuclei. Fluorescence intensity analysis was performed using confocal microscope.

Results were evaluated using statistical significance of Dunn’s multiple comparisons test. Blue bars represent average values and error bars represent standard error values.

1Mosser D. and Zhang X.: Activation of Murine Macrophages. Curr Protoc Immunol; 2008

2Gumireddy K.: Mitogen-Activated Protein Kinase Pathway Mediates DBP—maf-Induced Apoptosis in RAW 264.7

Macrophages. J Cell Biochem; 2003

RESULTS

1) Reactive Oxygen Species production upon treatment with Omnia Drops and its components

Fig. la. ROS production in BMDM cells after incubation with Omnia Drops and its components. CS – chondroitin sulfate, OA – oleic acid, PMA – ROS inducer (positive control).

Fig. lb. ROS production in BMDM cells after incubation with ethanol dissolved oleic acid. OA — oleic acid, E – ethanol, PMA – ROS inducer (positive control).

 Fig. 2a. ROS production in RAW 246.7 cells after incubation with Omnia Drops and its components. CS – chondroitin sulfate, OA – oleic acid, PMA – ROS inducer (positive control).

Fig. 2b. ROS production in RAW 246.7 cells after incubation with ethanol dissolved oleic acid. E – ethanol, PMA – ROS inducer (positive control).

Fig. 3. ROS production in BMDM cells after incubation with Cytonic protein. CP

Fig. 4. ROS production in RAW 246.7 cells after incubation with Cytonic protein. CP

NOTE

All results were compared to the control sample (buffer only or E 0.05%) that did not contain any of the tested substances, and that was marked as 100% of the fluorescent signal intensity (delineated as a red dashed line across the chart). PMA effect was included in each chart showing the level of fluorescent signal measured for standard ROS inducer.

Oleic acid is insoluble in water and was mixed with ethanol in order to increase its water solubility.

CONCLUSIONS

1. Treatment with Omnia resulted in increased oxidative stress caused by ROS generation in both BMDM and RAW 246.7 cells. Cells treated with Omnia diluted 10 and 100 times displayed higher fluorescence intensity in comparison to PMA (ROS inducer, positive control). Minor stimulation was observed in BMDM cells treated with 1000 times dilution (fluorescence intensity higher than negative control, lower than PMA). No effect was observed in RAW 246.7 cells treated with 1000 times diluted Omnia.

2. Treatment with particular components of Omnia revealed that chondroitin sulfate alone inhibited ROS generation, while oleic acid, both water and ethanol dissolved, stimulated ROS generation:

  • BMDM and RAW 246.7 cells treated with 2.5 and 0.25 mg/ml of chondroitin sulfate displayed lower fluorescence intensity in comparison to cells incubated only with buffer (negative control). No significant effect was observed for 0.025mg/m1concentration.
  • BMDM and RAW 246.7 cells treated with 1.25 and 0.125 mg/ml of oleic acid, both water and ethanol dissolved, displayed higher fluorescence intensity in comparison with PMA (ROS inducer, positive control). No significant effect was observed for 0.0125 mg/m1 concentration, except for RAW 246.7 cells treated with ethanol dissolved oleic acid, where stimulation was observed as well.

3. Treatment with Cytonic protein resulted in increased oxidative stress caused by ROS generation in both BMDM and RAW 246.7 cells. Both cell lines treated with Cytonic at 3.3 and 0.33 ug/ml concentrations displayed higher fluorescence intensity in comparison to PMA (ROS inducer, positive control). Cells treated with Cytonic at 0.033 ug/ml concentration displayed higher fluorescence intensity in comparison to control sample (buffer only), but lower when compared to PMA (ROS inducer, positive control) indicating minor stimulating effect.

4. To sum up, both Omnia Drops and Cytonic stimulate ROS generation in BMDM and RAW 246.7 macrophage cell lines, which may be translated into enhanced immune activity of the cells.

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